Variability in the pattern of random amplified polymorphic DNA

被引:26
作者
Khandka, DK
Tuna, M
Tal, M
Nejidat, A
Golan-Goldhirsh, A
机构
[1] Ben Gurion Univ Negev, Jacob Blaustein Inst Desert Res, Albert Katz Ctr Desert Agrobiol, Desert Plant Biotechnol Lab, IL-84990 Sede Boqer, Israel
[2] Ben Gurion Univ Negev, Dept Life Sci, IL-84105 Beer Sheva, Israel
关键词
amplification variability; polymerase chain reaction; random amplified polymorphic DNA; reaction composition;
D O I
10.1002/elps.1150181522
中图分类号
Q5 [生物化学];
学科分类号
071010 ; 081704 ;
摘要
The random amplified polymorphic DNA (RAPD) technique is a simple method to detect DNA polymorphism. It is sensitive to reaction conditions. Small changes in the reactants' concentration cause variations in amplification products. Using DNA from Asparagus officinalis, Dactylis glomerata, Mercurialis annua and Escherichia coli, we examined variability in the amplification pattern associated with reaction constituents. An increase in the ratio of Tag DNA polymerase to DNA in the reaction increased the number of amplified fragments. Increasing the concentration of primer resulted in the amplification of low molecular weight DNA fragments, while lowering: the concentration resulted in high molecular weight fragments. Subsets or amplified fragments required different concentrations of magnesium for their highest intensity. Mechanical shearing of DNA obtained by sonication led to reduction in amplification of a subset of products. Enzymatic fragmentation of DNA by restriction enzymes led to loss or gain of specific fragments, depending on the DNA, primer, and restriction enzyme. RAPD markers of pooled DNA of anonymous pedigree should be critically evaluated for frequent 'false positive' markers.
引用
收藏
页码:2852 / 2856
页数:5
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